Method for producing mercaptans by disulfide enzyme hydrogenolysis

ABSTRACT

Provided is an enzymatic process for preparing mercaptans from disulfides.

The present invention relates to a process for the production byenzymatic catalysis of mercaptans, in particular of methyl mercaptan,from disulfides, in particular dimethyl disulfides, and using organicreducing compounds.

Mercaptans are highly useful in very numerous fields, for example asflavourings, odorants for gases, chain transfer agents inpolymerisation, starting materials for the pharmaceutical or cosmeticindustry, for the synthesis of antioxidants, extreme-pressure oranti-wear additives for lubrication. These examples do not in any waylimit the uses of the mercaptans known at present and which can beprepared by virtue of the process of the invention.

In particular, the first of the mercaptans, methyl mercaptan (CH₃SH), isvery industrially beneficial, in particular as starting material in thesynthesis of methionine, an essential amino acid very widely used inanimal feed. Methyl mercaptan is also a starting material very widelyused for the synthesis of numerous other molecules.

Mercaptans may be synthesised by numerous methods such as thesulfhydration of alcohols, the catalytic or photochemical addition ofhydrogen sulfide onto unsaturated organic compounds, the substitution ofhalides, epoxides or organic carbonates by means of hydrogen sulfide,and others.

In particular, methyl mercaptan is currently produced industrially onthe tonne scale from methanol and hydrogen sulfide according to thereaction (1):

CH₃OH+H₂S→CH₃SH+H₂O  (1)

These processes have the drawbacks of requiring methanol (CH₃OH), ofsynthesising hydrogen sulfide (H₂S, from hydrogen and sulfur forexample, which also then requires the synthesis of hydrogen), and giverise to by-products of dimethyl ether (CH₃OCH₃), dimethyl sulfide(CH₃SCH₃) type, and products of cracking and water, which impliesnumerous steps for purification of the methyl mercaptan.

By way of examples, the description of processes based on thesereactions will be found in patent applications such as WO2013092129,WO2008118925, WO2007028708, WO2006015668 and WO2004096760.

It may prove economically advantageous (to avoid methanol synthesis) towish to produce methyl mercaptan from carbon monoxide, hydrogen andhydrogen sulfide, according to the following synthesis scheme (2):

CO+2H₂+H₂S→CH₃SH+H₂O  (2)

However, these processes have the drawbacks of requiring synthesis gas(CO/H₂) and therefore carrying out steam reforming of a source ofhydrocarbons, having the correct proportions between CO and H₂, hencebeing able to adjust the CO/H₂ ratio with what is referred to as the“water-gas shift reaction” (CO+H₂O→CO₂+H₂), and synthesising H₂S.

These processes also generally lead to large proportions of CO₂ asby-product, and also to methane, dimethyl sulfide and water. By way ofexample, descriptions of these processes will be found in patentapplications US2010286448, US2010094059, US2008293974, US2007213564.

Yet other processes have been described, and combine different reactionssuch as:

-   -   1) Formation of CS₂ and H₂S from methane and sulfur (3):

CH₄+4S→CS₂+2H₂S  (3)

-   -   2) Hydrogenation of CS₂ (4):

CS₂+3H₂→CH₃SH+H₂S  (4)

It is also possible to use the excess H₂S from reactions (3) and (4) inthe reaction with methanol (reaction 1) or the reaction with synthesisgas (reaction 2) to further give methyl mercaptan.

These processes obviously combine the drawbacks described for reactions(1) and (2) with the additional difficulty of having excess hydrogen tocarry out reaction (4). Descriptions of these processes will be found inpatent applications US2011015443, or, more specifically in relation toreaction (4), in application WO2010046607.

Application WO200196290 proposes a process for synthesising methylmercaptan directly from methane and H₂S with coproduction of hydrogen.This direct reaction between methane and H₂S occurs by means of a pulsedplasma with corona discharge. Since this application does not describeany examples of synthesis, it may appear difficult to imagine a processfor the large-scale industrial synthesis of methyl mercaptan with thistechnology. Moreover, this process requires the synthesis of H₂S if thelatter is not available.

For its part, patent application EP0649837 proposes a process for thesynthesis of methyl mercaptan by catalytic hydrogenolysis, withtransition metal sulfides, of dimethyl disulfide with hydrogen. Althoughthis process is efficient, it requires relatively high temperatures ofthe order of 200° C. to obtain industrially advantageous levels ofproductivity.

Those skilled in the art also know that it is possible to prepare methylmercaptan by acidification of an aqueous solution of sodium methylmercaptide (CH₃SNa). This method has the major drawback of producinglarge amounts of salts, such as sodium chloride or sodium sulfate,depending on whether hydrochloric acid or sulfuric acid is used. Thesesaline aqueous solutions are often very difficult to treat and thetraces of foul-smelling products which remain mean that this methodcannot be readily envisaged on the industrial scale.

The processes for synthesising mercaptans higher than methyl mercaptanalso have numerous drawbacks. Thus, the substitution of alcohols withhydrogen sulfide requires high temperatures, and often pressures, andleads to undesired by-products of olefin, ether and sulfide type.

The catalytic or photochemical addition of hydrogen sulfide ontounsaturated compounds often occurs under slightly milder conditions thanabove, but also leads to numerous by-products formed by isomerisation ofthe starting material, by non-regioselective addition or by doubleaddition which gives sulfides. Finally, the substitution of halogenatedderivatives gives rise to processes which generate large amounts ofeffluents and saline waste which are not easily reconcilable withindustrial processes.

The subject of the present invention is to propose a novel process forpreparing mercaptans, in particular methyl mercaptan, which does nothave the drawbacks described in the processes known from the prior artlaid out above.

More particularly, a first subject-matter of the present invention isthe process for the preparation of a mercaptan of formula R—SH,comprising at least the steps of:

-   -   a) preparation of a mixture comprising:        -   1) a disulfide of formula R—S—S—R′,        -   2) a catalytic amount of amino acid bearing a thiol group or            of a thiol-group-containing peptide,        -   3) a catalytic amount of an enzyme catalysing the reduction            of the disulfide bridge created between two equivalents of            said amino acid bearing a thiol group or of said            thiol-group-containing peptide,        -   4) a catalytic amount of an enzyme catalysing the            dehydrogenation of the organic reducing compound involved in            step b),        -   5) a catalytic amount of a cofactor common to the two            enzymes catalysing the reduction and the dehydrogenation,    -   b) addition of an organic reducing compound in a stoichiometric        amount relative to the disulfide of formula R—S—S—R′,    -   c) carrying out the enzymatic reaction,    -   d) recovery of the mercaptan of formula R—SH and of the        mercaptan of formula R′—SH,    -   e) separation and optional purification of the mercaptan of        formula R—SH and of the mercaptan of formula R′—SH.

Generally speaking, the enzyme catalysing the reduction of the disulfidebridge created between two equivalents of said amino acid bearing athiol group or said thiol-group-containing peptide is a reductaseenzyme. The term “reductase” is used in the remainder of the descriptionfor explaining the present invention. Similarly, the enzyme catalysingthe dehydrogenation of the organic reducing compound involved in step b)is generally referred to as a dehydrogenase enzyme, the term“dehydrogenase” being chosen in the remainder of the description forexplaining the present invention.

Among the cofactors common to the two enzymes catalysing the reductionand the dehydrogenation (reductase and dehydrogenase), mention may bemade, by way of non-limiting examples, of flavinic cofactors andnicotinic cofactors. Preference is given to using nicotinic cofactorsand more particularly nicotinamide adenine dinucleotide (NAD), or betterstill nicotinamide adenine dinucleotide phosphate (NADPH). The cofactorslisted above are advantageously used in their reduced forms (for exampleNADPH, H+) and/or their oxidized forms (for example NADP+), that is tosay that they may be added in these reduced and/or oxidized forms intothe reaction medium.

In one embodiment of the invention, the amino acid bearing a thiol groupand/or the thiol-group-containing peptide may be in the form of thedisulfide of said amino acid and/or of said peptide, respectively, forexample glutathione in the form of glutathione disulfide.

The organisation and the order of the additions of the differentcomponents of steps a) and b) of the process defined above may becarried out in different ways. In any case, the enzymatic reaction ofstep c) is triggered by the addition of one of the components of thecatalytic system: either an enzyme, or one of the compounds added in astoichiometric amount (disulfide or organic reducing compound), or oneof the compounds added in a catalytic amount (amino acid bearing a thiolgroup or thiol-group-containing peptide or disulfide corresponding tosaid molecules or else the cofactor).

Even more particularly, a subject-matter of the present invention is theprocess for the preparation of a mercaptan of formula R—SH, comprisingat least the steps of:

-   -   a′) preparation of a mixture comprising:        -   a disulfide of formula R—S—S—R′,        -   a catalytic amount of amino acid bearing a thiol group or of            a thiol-group-containing peptide,        -   a catalytic amount of reductase enzyme corresponding to said            amino acid bearing a thiol group or to said            thiol-group-containing peptide,        -   a catalytic amount of NADPH,    -   b′) addition of an organic reducing compound in a stoichiometric        amount relative to the disulfide and DMDS) with a catalytic        amount of the corresponding dehydrogenase enzyme,    -   c′) carrying out the enzymatic reaction,    -   d′) recovery of the mercaptan of formula R—SH and of the        mercaptan of formula R′—SH,    -   e′) separation and optional purification of the mercaptan of        formula R—SH and of the mercaptan of formula R′—SH.

Within the context of the present invention, any disulfide correspondingto the general formula R—S—S—R′ may be involved in the process forproducing mercaptan. In the general formula R—S—S—R′, R and R′, whichare identical or different, represent independently of one another alinear, branched or cyclic hydrocarbon-based radical comprising from 1to 20 carbon atoms, said chain being saturated or bearing one or moreunsaturations in the form of double or triple bond(s). R and R′ may alsoform together, and with the sulfur atoms bearing them, a cyclic moleculecomprising from 4 to 22 atoms, preferably from 5 to 10 atoms.

According to a preferred aspect, the radicals R and R′, which areidentical or different, are chosen independently of one another fromlinear or branched, saturated or unsaturated alkyl, cycloalkyl, aryl,alkylaryl or arylalkyl radicals comprising from 1 to 20 carbon atoms,preferably from 1 to 12 carbon atoms, more preferably still from 1 to 6carbon atoms and optionally functionalised by one or more functionschosen, nonlimitingly and by way of example, from alcohol, aldehyde,ketone, acid, amide, nitrile or ester functions or else functionsbearing sulfur, phosphorus, silicon or halogen.

The disulfide of formula R—S—S—R′ is able to be reduced, according tothe process of the invention, to mercaptan of formula R—SH and mercaptanof formula R′—SH. When R is different to R′, reference is made toasymmetrical disulfides, and when R and R′ are identical, reference ismade to symmetrical disulfides. In the case of symmetrical disulfidesR—S—S—R, the process of the invention leads to a mercaptan of formulaR—SH. According to a particularly preferred aspect of the invention,dimethyl disulfide (DMDS) is used with the aim of producing methylmercaptan CH₃SH.

In the case of asymmetrical disulfides R—S—S—R′, the process of theinvention leads to a mixture of mercaptans of formulae R—SH and R′—SH,which may either be used as is or else subjected to one or moreseparation operations well known to those skilled in the art, forexample distillation.

It is also possible to use, in the process of the invention, mixtures ofone or more symmetrical and/or asymmetrical disulfides. Possiblemixtures of disulfides may comprise DSOs (disulfide oils), said DSOsthus finding a highly advantageous possibility of exploitation.

According to the process of the invention, the mercaptan(s) produced aregenerally recovered in the form of a solid, a liquid and/or a gas.

The production process according to the invention is based on theenzymatic reduction of disulfides, in particular dimethyl disulfide,with an organic reducing compound, which is a hydrogen donor as will bedefined below, according to the following reaction, illustrated withdimethyl disulfide leading to methyl mercaptan, using glucose as organicreducing compound (hydrogen donor):

It has now been discovered that this reaction is readily catalysed bythe enzymatic system employing a thiol-group-containing amino acid or athiol-group-containing peptide, for example glutathione, in the form ofan (amino acid or peptide)/corresponding reductase enzyme complex,regenerated by the hydrogen-donating organic compound, as described inthe appended FIG. 1.

Thus, according to the illustration in FIG. 1, the peptide (the examplerepresented being glutathione) reduces the disulfide (DMDS represented)to mercaptan (methyl mercaptan represented) by converting into a peptidewith a disulfide bridge (glutathione disulfide represented). Thereductase enzyme (glutathione reductase represented, enzymeclassification numbers EC 1.8.1.7 or EC 1.6.4.2) regenerates the peptide(glutathione) and this same enzyme is regenerated by a redox enzymaticcomplex well known to those skilled in the art, for example theNADPH/NADP+ (nicotinamide adenine dinucleotide phosphate (reduced formand oxidized form)) complex. NADP+ is in turn regenerated to NADPH bymeans of the dehydrogenase enzyme corresponding to the organic reducingcompound used (here, glucose dehydrogenase, EC 1.1.1.47) by virtue ofsaid organic reducing compound (glucose represented) which provideshydrogen (hydrogen donor) by converting to its oxidized form (here,gluconolactone).

In other words, the enzyme catalysing the reaction (glutathionereductase represented with the example enzyme classification numbers EC1.8.1.7 or EC 1.6.4.2) regenerates the peptide (glutathione) whileoxidizing the cofactor (NADPH,H+ represented). The oxidized form (NADP+represented) is then reduced by means of a “recycling” redox enzymaticcomplex well known to those skilled in the art and comprising thedehydrogenase enzyme involved (glucose dehydrogenase represented withthe example enzyme classification number EC 1.1.1.47) and the organicreducing molecule (glucose represented). The oxidized form of theorganic reducing compound is then obtained (gluconolactone represented).

According to a most particularly suited embodiment, theglutathione/glutathione disulfide system combined with the glutathionereductase enzyme makes it possible according to the present invention toreduced the DMDS to methyl mercaptan.

Glutathione is a tripeptide widely used in biology. In reduced form(glutathione) or oxidized form (glutathione disulfide), this speciesforms an important redox couple in cells. Thus, glutathione is vital foreliminating heavy metals from organisms. Thus, for example, applicationWO05107723 describes a formulation in which glutathione is used to forma chelating preparation and patent U.S. Pat. No. 4,657,856 teaches thatglutathione also makes it possible to break down peroxides such as H₂O₂into H₂O via glutathione peroxidase. Finally, glutathione also makes itpossible to reduce disulfide bridges present in proteins (RonaChandrawati, “Triggered Cargo Release by Encapsulated EnzymaticCatalysis in Capsosomes”, Nano Lett., (2011), vol. 11, 4958-4963).

According to the process of the invention, a catalytic amount of aminoacid bearing a thiol group or of a thiol-group-containing peptide isused to produce mercaptans from disulfides.

Among the amino acids bearing a thiol group which may be used in theprocess of the present invention, mention may be made by way ofnonlimiting examples of cysteine and homocysteine. In these cases, theredox enzymatic systems used which can regenerate the catalytic cycle inthe same way as the system cysteine/cystine reductase EC 1.8.1.6 andhomocysteine/homocysteine reductase.

Among the peptides bearing a thiol group which may be used in theprocess of the present invention, mention may be made by way ofnonlimiting examples of glutathione and thioredoxin. Theglutathione/glutathione reductase system described above may thus bereplaced by the thioredoxin (CAS No. 52500-60-4)/thioredoxin reductase(EC 1.8.1.9 or EC 1.6.4.5) system.

Glutathione and the glutathione/glutathione reductase system are mostparticularly preferred for the present invention, due to the costs ofthese compounds and the ease with which they are procured.

Among the organic reducing compounds which may be used within thecontext of the present invention, hydrogen-donating compounds are mostparticularly preferred, and among these, the entirely suitable compoundsare hydrogen-donating organic reducing compounds bearing a hydroxylfunction, such as alcohols, polyols, sugars, etc.

The enzyme used is an enzyme able to dehydrogenate the hydrogen-bearingcompound, for example an alcohol dehydrogenase. Glucose is a mostparticularly well-suited sugar to be used in the process of the presentinvention with glucose dehydrogenase to give gluconolactone.

In the process according to the invention, only the disulfide(s) and theglucose are used in a stoichiometric amount and all the other components(amino acid or peptide, cofactor (for example NADPH) and the 2 enzymes)are used in catalytic amounts.

The advantages brought about by the process of the invention arenumerous. Among these advantages, mention may be made of the possibilityof working in aqueous or aqueous-organic solution, under very mildtemperature and pressure conditions and under pH conditions close toneutrality. All these conditions are typical of a “green” or“sustainable” biocatalytic process.

Another advantage when the process uses dimethyl disulfide is that themethyl mercaptan produced, which is in the gaseous state under thereaction conditions, leaves the reaction medium as it is formed. Themethyl mercaptan may therefore be directly used, upon leaving thereactor, in an application further downstream. It can also be readilyliquefied cryogenically for example, if it is desired to isolate it. Itis optionally possible to accelerate its departure from the reactionmedium by introducing a low flow rate of nitrogen, by bubbling.

The dimethyl disulfide (DMDS) may be produced at another site frommethyl mercaptan and an oxidizer such as oxygen, sulfur or aqueoushydrogen peroxide solution, for example, or else from dimethyl sulfateand sodium disulfide. The DMDS may also originate from a source ofdisulfide oils (DSO), as indicated above, then be purified for exampleby reactive distillation as described in application WO2014033399. Itshould be noted that the DSOs may also be used as is, without thenecessity for purification between the different disulfides composingthem. A mixture of mercaptans is then obtained by applying the processof the invention.

When DMDS is used as disulfide, the process according to the inventionis can then be considered as a process which makes it possible to avoidtransporting methyl mercaptan from its site of production by existingindustrial routes, to its site of use, if they are different. Indeed,methyl mercaptan is a toxic and extremely foul-smelling gas at roomtemperature, which significantly complicates its transportation, whichis already heavily regulated unlike DMDS. The process described in thepresent invention can therefore be used to produce methyl mercaptandirectly on the site of use of the latter.

Since the DMDS is consumed in the reaction and the methyl mercaptanleaves the reaction medium as it is formed, only the product of thedehydrogenation of the organic reducing compound, for examplegluconolactone, accumulates in the reaction medium, if it is assumedthat glucose and DMDS are fed continuously. When the gluconolactoneconcentration exceeds the saturation point under the reactionconditions, it will precipitate out and may then be isolated from thereaction medium by any means known to those skilled in the art.

Gluconolactone may have several uses. It is for example used as a foodadditive, known by the reference E575. Gluconolactone is hydrolysed inacidic aqueous media to form gluconic acid, also used as a food additive(E574). Gluconolactone is also used for the production of tofu (cf.CN103053703) for the food industry.

Especially and advantageously, in the sense that it represents the“waste” from the process according to the present invention,gluconolactone may replace glucose in a possible fermentation reactionto produce either bioethanol or any other molecule originating from thefermentation of sugar or starch.

Indeed, certain bacteria may use gluconolactone as carbon source infermentation, as described by J. P. van Dijken, “Novel pathway foralcoholic fermentation of gluconolactone in the yeast Saccharomycesbulderi”, J. Bacteriol., (2002), Vol. 184(3), 672-678.

Yet other sugars may be used in the process of the invention, and forexample it is possible to replace the glucose/gluconolactone/glucosedehydrogenase system with the following system: glucose6-phosphate/6-phosphoglucono-δ-lactone/glucose6-phosphate dehydrogenase(EC 1.1.1.49).

It is also possible, in the process of the invention, to use an alcoholin place of the sugar, and thus to use the following general systeminstead of the glucose/gluconolactone/glucose dehydrogenase system:alcohol/ketone or aldehyde/alcohol dehydrogenase (EC 1.1.1) and moreparticularly the isopropanol/acetone/isopropanol dehydrogenase system(EC 1.1.1.80).

Indeed, this system makes it possible to obtain, when DMDS is used asdisulfide, a mixture consisting of methyl mercaptan (MeSH) and acetonewhich leaves the reaction medium (therefore no accumulation of anyproduct). The MeSH and the acetone may be easily separated by simpledistillation if desired. In the case of other disulfides, depending onthe boiling point of the mercaptan formed and its solubility in thereaction medium, the acetone may be readily removed from the medium andthe mercaptan may optionally settle out of the reaction medium, in orderto be easily separated.

Generally, the reaction temperature is within a range extending from 10°C. to 50° C., preferably between 15° C. and 45° C., more preferablybetween 20° C. and 40° C.

The pH of the reaction may be between 6 and 8, preferably between 6.5and 7.5. The pH of the reaction medium may be adjusted by means of abuffer. Entirely preferably, the pH of the phosphate buffer will bechosen to be 7.3.

The pressure used for the reaction may range from a reduced pressurecompared to atmospheric pressure to several bar (several hundred kPa),depending on the reagents and equipment used. In the case where DMDS isused as disulfide, a reduced pressure may indeed enable quickerdegassing of the methyl mercaptan formed, but has the drawback ofincreasing the saturated vapour pressures of the water and the DMDS,polluting the methyl mercaptan formed slightly more. Preferably, usewill be made of a pressure ranging from atmospheric pressure to 20 bar(2 MPa) and even more preferably the process will be carried out under apressure ranging from atmospheric pressure to 3 bar (300 kPa).

The process according to the invention can be carried out batchwise orcontinuously, in a glass or metal reactor depending on the operatingconditions selected and the reagents used.

The ideal organic reducing compound/disulfide molar ratio isstoichiometry (molar ratio=1) but may vary from 0.01 to 100, if thoseskilled in the art find any benefit therein, such as continuous additionof disulfide while the reducing compound is introduced from the startinto the reactor. Preferably, this molar ratio is chosen between 0.5 and5 overall, over the whole of the reaction.

The elements present in catalytic amounts in the mixture prepared instep a) above (amino acid bearing a thiol group or athiol-group-containing peptide, reductase enzyme, cofactor such as, forexample, NADPH) are easily available commercially or can be preparedaccording to techniques well known to those skilled in the art. Thesedifferent elements may be in solid or liquid form and may veryadvantageously be dissolved in water to be used in the process of theinvention. The enzymes used may also be grafted onto a support (in thecase of supported enzymes).

The aqueous solution of enzymatic complex comprising the amino acid orthe peptide may also be reconstituted by methods known to those skilledin the art, for example by permeabilization of cells which contain theseelements. This aqueous solution, a composition of which is given in thefollowing Example 1, may be used in contents by weight of between 0.01%and 20% relative to the total weight of the reaction medium. Preferably,a content of between 0.5% and 10% will be used.

According to another aspect, the present invention relates to the use ofan aqueous solution of enzymatic complex comprising an amino acidbearing a thiol function as defined above or a peptide bearing a thiolfunction as defined above, for the synthesis of a mercaptan from adisulfide.

The mixture which can be used for step a) of the process described aboveand comprising:

-   -   1) a disulfide of formula R—S—S—R′,    -   2) a catalytic amount of amino acid bearing a thiol group or a        thiol-group-containing peptide,    -   3) a catalytic amount of an enzyme catalysing the reduction of        the disulfide bridge created between two equivalents of said        amino acid bearing a thiol group or to said        thiol-group-containing peptide,    -   4) optionally a catalytic amount of an enzyme catalysing the        dehydrogenation of an organic reducing compound,    -   5) a catalytic amount of a cofactor common to the two enzymes        catalysing the reduction and the dehydrogenation,        where R and R′ are as defined above,        is novel and hence is part of the present invention.

In one embodiment of the invention, the amino acid bearing a thiol groupand/or the peptide bearing a thiol group can be in the form of thedisulfide of said amino acid and/or of said peptide, respectively.

More particularly, said mixture comprises:

-   -   a disulfide of formula R—S—S—R′,    -   a catalytic amount of amino acid bearing a thiol group or a        thiol-group-containing peptide,    -   a catalytic amount of reductase enzyme corresponding to said        amino acid bearing a thiol group or to said        thiol-group-containing peptide, and    -   a catalytic amount of NADPH,        where R and R′ are as defined above.

The invention will be better understood with the following examplesnonlimiting relative to the scope of the invention.

EXAMPLE 1

10 ml of glutathione enzymatic complex (Aldrich) and 19.2 g (0.1 mol) ofglucose are introduced into a reactor containing 150 ml of 0.1 mol/lphosphate buffer at pH 7.30. The solution of enzymatic complex contains:185 mg (0.6 mmol) of glutathione, 200 U of glutathione reductase, 50 mg(0.06 mmol) of NADPH and 200 U of glucose dehydrogenase. The reactionmedium is brought to 25° C. with mechanical stirring. A first sample istaken at t=0. Subsequently, the dimethyl disulfide (9.4 g, 0.1 mol) isplaced in a burette and added dropwise to the reactor; the reactionbegins. A stream of nitrogen is placed in the reactor. Gaschromatography analysis of the gases leaving the reactor shows virtuallyessentially the presence of nitrogen and methyl mercaptan (some tracesof water). These outlet gases are trapped in 20% sodium hydroxide inwater. The DMDS is introduced in 6 hours and the reaction is monitoredby potentiometric argentometric titration of the methyl mercaptan sodiumsalt in the trap at the outlet of the reactor. In addition, a final gaschromatography analysis of the reaction medium confirms the absence ofDMDS, and by UPLC/mass spectrometry traces of glucose and the virtuallyexclusive presence of gluconolactone are found.

EXAMPLE 2

To the reaction medium of Example 1, 19.2 g (0.1 mol) of glucose arereintroduced in one go, and 9.4 g (0.1 mol) of DMDS are reintroduceddropwise in 6 hours. The reaction is monitored in the same way as inExample 1, after having changed the 20% sodium hydroxide solution at theoutlet of the reactor. The analyses at the end of the reaction confirmthe complete disappearance of the DMDS, totally converted into methylmercaptan found in sodium salt form in the sodium hydroxide solution.Only the gluconolactone is analysed and found in the reaction medium atthe end of the reaction. This example shows the robustness of thecatalytic system through its reproducibility.

1.-13. (canceled)
 14. A mixture, comprising: a disulfide of formulaR—S—S—R′, wherein R and R′, independently, represent a linear, branchedor cyclic hydrocarbon-based radical comprising from 1 to 20 carbonatoms, wherein the hydrocarbon-based radical is saturated or containsone or more unsaturations in the form of double or triple bond(s), or Rand R′ form together, and with the sulfur atoms bearing them, a cyclicgroup comprising from 4 to 22 atoms, a catalytic amount of amino acidbearing a thiol group or of a thiol-group-containing peptide, whereinthe amino acid bearing a thiol group or the thiol-group-containingpeptide may optionally be in the form of the corresponding disulfide, acatalytic amount of an enzyme catalyzing the reduction of a disulfidebridge created between two equivalents of the amino acid bearing a thiolgroup or the thiol-group-containing peptide, optionally, a catalyticamount of an enzyme catalyzing the dehydrogenation of an organicreducing compound, and a catalytic amount of a cofactor common to thetwo enzymes catalyzing the reduction and the dehydrogenation.
 15. Themixture of claim 14, comprising: a disulfide of formula R—S—S—R′, acatalytic amount of the amino acid bearing a thiol group or thethiol-group-containing peptide, a catalytic amount of a reductase enzymecorresponding to the amino acid bearing a thiol group or thethiol-group-containing peptide, and a catalytic amount of NADPH.
 16. Themixture of claim 14, wherein the disulfide of formula R—S—S—R′ isdimethyl disulphide.
 17. The mixture of claim 14, wherein the amino acidbearing a thiol group or the peptide bearing a thiol group is chosenfrom cysteine, homocysteine, glutathione and thioredoxin.
 18. Themixture of claim 14, wherein the organic reducing compound is ahydrogen-donating organic reducing compound bearing a hydroxyl function,chosen from alcohols, polyols, sugars, etc.
 19. The mixture of claim 14,wherein the organic reducing compound is chosen from glucose, glucose6-phosphate and isopropanol.
 20. The mixture of claim 14, wherein theorganic reducing compound/disulfide molar ratio is between 0.01 and 100.21. The mixture of claim 14, wherein the organic reducingcompound/disulfide molar ratio is between 0.5 and
 5. 22. The mixture ofclaim 14, wherein: the disulfide of formula R—S—S—R′ is dimethyldisulfide (DMDS), the amino acid bearing a thiol group or the peptidebearing a thiol group is chosen from cysteine, homocysteine, glutathioneand thioredoxin, the organic reducing compound is a hydrogen-donatingorganic reducing compound bearing a hydroxyl function and is chosen fromalcohols, polyols, and sugars, and the cofactor is a flavinic cofactoror a nicotinic cofactor.
 23. The mixture of claim 14, wherein: thedisulfide of formula R—S—S—R′ is dimethyl disulfide (DMDS), the aminoacid bearing a thiol group or the peptide bearing a thiol group isglutathione, the organic reducing compound is glucose, and the cofactoris NADPH.